THE MECHANISM OF NATURAL OCCURRENCE OF 2 CLOSELY LINKED HBV PRECORE PREDOMINANT MUTATIONS

Two precore predominant mutations of human hepatitis B virus (HBV) at either nucleotide (nt) 1896 or nt 1899 often occur in combination. At nt 1896, a G to A mutation creates a TAG stop codon at codon 28 of pre core protein. At nt 1899, a G to A mutation changes glycine at codon 2 9 to aspartic acid. To assess the effect of each individual mutation a s well as any interaction between these two mutations, HBV derivatives bearing one or both precore predominant mutations have been construct ed. HBV e-Ag-negative mutants bearing a TAG stop codon mutation at cod on 28 uniformly replicate at least 20-fold better than mutants bearing a TGA stop codon at the same amino acid position, irrespective of the sequence context at nt 1899. A single mutation at nt 1899, changing t he wild-type G to a pyrimidine (T or C) is deleterious to viral RNA en capsidation and DNA replication, Our results explain in part why only a purine (G or A) at nt 1899, never a pyrimidine, is observed in natur al HBV genomes. The effects caused by these two closely linked mutatio ns on viral replication are not independent of each other. The stringe nt selection for a highly efficient RNA encapsidation element may play a crucial role in the natural occurrence of these two closely linked precore mutations. The putative 27-amino-acid peptide resulting from t he truncation of precore by the nt 1896 mutation has no apparent effec t on viral replication. The preferential occurrence of the G to A muta tion at nt 1896 and 1899, instead of at other nonpredominant positions , is likely to be a combined consequence of both selection and higher intrinsic mutation frequency at these positions. (C) 1995 Academic Pre ss, Inc.